Determination of glycan structure from tandem mass spectra

Glycans are molecules made from simple sugars that form complex tree structures. Glycans constitute one of the most important protein modifications and identification of glycans remains a pressing problem in biology. Unfortunately, the structure of glycans is hard to predict from the genome sequence of an organism. In this paper, we consider the problem of deriving the topology of a glycan solely from tandem mass spectrometry (MS) data. We study, how to generate glycan tree candidates that sufficiently match the sample mass spectrum, avoiding the combinatorial explosion of glycan structures. Unfortunately, the resulting problem is known to be computationally hard. We present an efficient exact algorithm for this problem based on fixed-parameter algorithmics that can process a spectrum in a matter of seconds. We also report some preliminary results of our method on experimental data, combining it with a preliminary candidate evaluation scheme. We show that our approach is fast in applications, and that we can reach very well de novo identification results. Finally, we show how to count the number of glycan topologies for a fixed size or a fixed mass. We generalize this result to count the number of (labeled) trees with bounded out degree, improving on results obtained using Pólya's enumeration theorem.     

Linking the evolution of habitat choice to ecosystem functioning: direct and indirect effects of pond-reproducing fire salamanders on aquatic-terrestrial subsidies

Shifts in life history traits and in the behaviour of species can potentially alter ecosystem functioning. The reproduction of the central European fire salamander (Salamandra salamandra), which usually deposits its larvae in first-order streams, in small pool and pond-like habitats, is an example of a recent local adaptation in this species. Here we aimed to quantify the direct and indirect effects of the predatory larvae on the aquatic food webs in the ponds and on the flux of matter between the ponds and adjacent terrestrial habitats. Our estimates are based on biomass data of the present pond fauna as well as on the analysis of stomach content data, growth rates and population dynamics of the salamander larvae in pond habitats. By their deposition of larvae in early spring, female fire salamanders import between 0.07 and 2.86 g dry mass m^?2 larval biomass into the ponds. Due to high mortality rates in the larval phase and the relatively small size at metamorphosis of the pond-adapted salamanders compared to stream-adapted ones, the biomass export of the metamorphosed salamanders clearly falls below the initial biomass import. Catastrophic events such as high water temperatures and low oxygen levels may even occasionally result in mass mortalities of salamander larvae and thus in a net 100 % import of the salamander biomass into the pond food webs. Indirect effects further accelerate this net import of matter into the aquatic habitat, e.g. the feeding of salamanders on aquatic insect larvae with the emergence of terrestrial adults—thus preventing export—and on terrestrial organisms that fall on the water surface (supporting import). This study demonstrates that the adaptation of salamanders to pond reproduction can alter food web linkages across ecosystem boundaries by enhancing the flux of materials and energy from terrestrial (i.e. forest) to the aquatic (i.e. pond) habitat.     

Biogeography and phylogenetic diversity of a cluster of exclusively marine myxobacteria

Myxobacteria are common in terrestrial habitats and well known for their formation of fruiting bodies and production of secondary metabolites. We studied a cluster of myxobacteria consisting only of sequences of marine origin (marine myxobacteria cluster, MMC) in sediments of the North Sea. Using a specific PCR, MMC sequences were detected in North Sea sediments down to 2.2 m depth, but not in the limnetic section of the Weser estuary and other freshwater habitats. In the water column, this cluster was only detected on aggregates up to a few meters above the sediment surface, but never in the fraction of free-living bacteria. A quantitative real-time PCR approach revealed that the MMC constituted up to 13% of total bacterial 16S rRNA genes in surface sediments of the North Sea. In a global survey, including sediments from the Mediterranean Sea, the Atlantic, Pacific and Indian Ocean and various climatic regions, the MMC was detected in most samples and to a water depth of 4300 m. Two fosmids of a library from sediment of the southern North Sea containing 16S rRNA genes affiliated with the MMC were sequenced. Both fosmids have a single unlinked 16S rRNA gene and no complete rRNA operon as found in most bacteria. No synteny to other myxobacterial genomes was found. The highest numbers of orthologues for both fosmids were assigned to Sorangium cellulosum and Haliangium ochraceum. Our results show that the MMC is an important and widely distributed but largely unknown component of marine sediment-associated bacterial communities.     

Supersize me—new insights into cortical expansion and gyration of the mammalian brain

EMBO J 32 13, 1817–1828 doi:10.1038/emboj.2013.96; published online April262013During evolution, the mammalian brain massively expanded its size. However, the exact roles of distinct neural precursors, identified in the developing cortex during embryogenesis, for size expansion and surface folding (i.e., gyration) remain largely unknown. New findings by Nonaka-Kinoshita et al advance our understanding of embryonic neural precursor function by identifying cell type-selective functions for size expansion and folding, and challenge previously held concepts of mammalian brain development.Over the course of evolution, the mammalian brain massively expanded its size and complexity, which is believed to be responsible for an increase in cognitive functions and intellectual skills. The increase in brain size and number of cortical neurons is primarily due to an increased surface area by generating folds (gyrations) while the cortical thickness remained relatively constant (Lui et al, 2011). In the last decade, substantial progress has been made in identifying the cellular sources of cortex development. Using genetic lineage tracing of individual cell populations and time-lapse imaging of rodent and human slices of the embryonic cortex, radial glial cells (RGCs) were identified as the primary progenitors or neural stem cells (NSCs) in the developing cortex (Gotz and Huttner, 2005). Simplified, RG in the ventricular zone (VZ) line the ventricular surface and self-renew through symmetric divisions or give rise to basal progenitors (BPs; also called intermediate progenitors) in the subventricular zone (SVZ) that typically divide symmetrically and generate neurons. In contrast to the lissencephalic rodent brain, the developing cortex of gyrated mammals (e.g., humans and ferrets) contains a large number of basal radial glial (bRG) cells that reside in the outer subventricular zone (OSVZ), retain a cellular process that is connected to the pial surface and that are, in contrast to BPs, multipotent, meaning that they have the potency to generate diverse neural cell types (Fietz et al, 2010; Hansen et al, 2010; Reillo et al, 2011).Largely based on the anatomical differences between the developing cortex of lissencephalic and gyrencephalic brains, several hypotheses have been formulated aiming to explain the massive increase in size and induction of brain folding during mammalian evolution. One prominent hypothesis, called the radial unit hypothesis, suggests that the expansion of RGCs lining the ventricle leads to an increase of radial units that generate neurons and thus is responsible for the increase of surface area (Rakic, 1995). Others proposed that the increase in size and folding could be due to an increase in BP expansion in the SVZ compared to RGC numbers in the VZ, a hypothesis called the intermediate progenitor model (Kriegstein et al, 2006). These hypotheses were helpful to start explaining mammalian brain evolution, but testing the exact role of different neural precursors remained extremely challenging due to technical difficulties to selectively manipulating the proliferative activity of distinct precursor populations. Even though previous approaches were successful in enhancing brain size/neuron numbers in mouse models (e.g., by ectopically enhancing WNT signalling activity or manipulating the activity of the small RhoGTPase Cdc42 in neural precursors), these strategies had the drawback that the normal six-layered cortical topography was disrupted, making it difficult to draw definite conclusions (Chenn and Walsh, 2002; Cappello et al, 2006).In a collaborative work from the Calegari and Borrell laboratories, Nonaka-Kinoshita et al, 2013 now used an elegant approach to selectively enhance proliferation of distinct precursor populations in the mouse and ferret developing cortex. They used a previously described approach manipulating cell cycle length and subsequently proliferation by overexpressing the cell cycle regulators cdk4 and cyclinD1 that is sufficient to enhance neurogenesis without affecting cortical layering (a system called 4D) (Lange et al, 2009). For their mouse experiments, Nonaka-Kinoshita et al used a transgenic strategy to transiently overexpress 4D in nestin-expressing precursors using a tetracycline-controlled gene expression system (nestin^rtTA/tetbi^4D). With this approach, they selectively enhanced proliferation of BPs in the SVZ without affecting the number or proliferation of RGCs in the VZ (Nonaka-Kinoshita et al, 2013). Strikingly, targeted expansion of BPs induced a substantial increase in surface area but was not sufficient to induce cortical folding in the otherwise smooth mouse cortex, challenging the radial unit hypothesis and the intermediate progenitor model with regard to their predictions on the effects on size and/or gyration of the cortex upon expansion of the BP pool. Complementing their findings of BP expansion in the lissencephalic mouse brain, Nonaka-Kinoshita et al used retroviral vectors and electroporation of 4D expression constructs to target 4D expression to neural precursors in the developing ferret cortex that is gyrated under physiological conditions. In the ferret, 4D expression induced proliferation of multipotent bRG located in the OSVZ, as outlined above, a cell type that is found predominantly in gyrated cortices compared to lissencephalic brains. Notably, enhanced proliferation of bRG triggered the formation of novel cortical folds, suggesting that indeed the expansion of bRG may represent a key event during evolution to induce gyration and subsequent surface expansion of the mammalian brain (Borrell and Reillo, 2012; Nonaka-Kinoshita et al, 2013) (Figure 1). This now experimentally supported hypothesis is strongly reinforced by two recent publications: one from (Tuoc et al, 2013) who found that deletion of the chromatin remodelling protein BAF170 increases the BP pool and subsequently enhances brain size; and another one from the Götz laboratory where it was found that experimentally reduced expression levels of the DNA-associated protein Trnp1 substantially increased the expansion of bRG and BPs, inducing folding of the normally lissencephalic mouse brain (Stahl et al, 2013). Taken together, these studies suggest that bRG in the OSVZ play an important role in cortical folding by enhancing the generation of neurons and by providing a glial scaffold for newborn neurons to disperse more laterally and thus to form folds in the developing brain (Reillo et al, 2011).Open in a separate windowFigure 1How different neural precursors appear to regulate size expansion and folding during mammalian brain development. (A) Shown are the main cellular components of the cortex of the lissencephalic mouse brain during embryonic development with RGCs (blue) lining the lateral ventricles in the VZ that generate BPs (yellow) in the SVZ and provide a scaffold for migrating neurons (left; green). Note that the mouse developing brain contains only a few bRG in the OSVZ (red). Notably, expansion of BPs using the 4D strategy developed in the Calegari laboratory increases surface area of the murine cortex without inducing the folding of the smooth mouse brain surface (right panel). (B) In contrast to lissencephalic animals, the developing cortices of species with gyrated brains (e.g., humans and ferrets) contain a substantial number of bRG located in the OSVZ (left panel). 4D-based, virus-mediated expansion of bRG in the ferret cortex leads to the induction of additional folds in the ferret cortex, indicating that the proliferative activity of bRG is critically involved in the extent of folding in physiologically gyrated brains (right panel).Even though this new study challenges previously held concepts regarding size expansion and folding of the mammalian brain, future studies are required that even more selectively enhance the proliferation and expansion of distinct precursor subtypes with high temporal and spatial control. Thus, the combination of sophisticated genetic tools to enhance precursor activity with detailed molecular analyses (e.g., analysing gene expression in highly folded versus unfolded brain regions, an approach that already showed differential levels of Trnp1 expression; Stahl et al, 2013) and live-imaging studies in the developing mammalian cortex will further enhance the understanding how our brains developed during evolution.     

miRNA-197 and miRNA-223 Predict Cardiovascular Death in a Cohort of Patients with Symptomatic Coronary Artery Disease

Background

Circulating microRNAs (miRNAs) have been described as potential diagnostic biomarkers in cardiovascular disease and in particular, coronary artery disease (CAD). Few studies were undertaken to perform analyses with regard to risk stratification of future cardiovascular events. miR-126, miR-197 and miR-223 are involved in endovascular inflammation and platelet activation and have been described as biomarkers in the diagnosis of CAD. They were identified in a prospective study in relation to future myocardial infarction.

Objectives

The aim of our study was to further evaluate the prognostic value of these miRNAs in a large prospective cohort of patients with documented CAD.

Methods

Levels of miR-126, miR-197 and miR-223 were evaluated in serum samples of 873 CAD patients with respect to the endpoint cardiovascular death. miRNA quantification was performed using real time polymerase chain reaction (RT-qPCR).

Results

The median follow-up period was 4 years (IQR 2.78–5.04). The median age of all patients was 64 years (IQR 57–69) with 80.2% males. 38.9% of the patients presented with acute coronary syndrome (ACS), 61.1% were diagnosed with stable angina pectoris (SAP). Elevated levels of miRNA-197 and miRNA-223 reliably predicted future cardiovascular death in the overall group (miRNA-197: hazard ratio (HR) 1.77 per one standard deviation (SD) increase (95% confidence interval (CI) 1.20; 2.60), p = 0.004, C-index 0.78; miRNA-223: HR 2.23 per one SD increase (1.20; 4.14), p = 0.011, C-index 0.80). In ACS patients the prognostic power of both miRNAs was even higher (miRNA-197: HR 2.24 per one SD increase (1.25; 4.01), p = 0.006, C-index 0.89); miRA-223: HR 4.94 per one SD increase (1.42; 17.20), p = 0.012, C-index 0.89).

Conclusion

Serum-derived circulating miRNA-197 and miRNA-223 were identified as predictors for cardiovascular death in a large patient cohort with CAD. These results reinforce the assumption that circulating miRNAs are promising biomarkers with prognostic value with respect to future cardiovascular events.     

Critical Evaluation of Phosphate Solubilizing Pseudomonads Isolated from a Partially Recultivated Potash Tailings Pile

Ten phosphate solubilizing pseudomonads isolated from a partially recultivated potash tailings pile in Germany were characterized and tested for their potential to assist in the ongoing recultivation process. Despite fertilization, the plants which are grown for recultivation show phosphate deficiency symptoms, and therefore the isolates are intended to be used as biofertilizer inoculants. On agar plates incubated at five different temperatures, some of the strains showed a temperature-dependent ability to solubilize tricalcium phosphate, while others performed the same at any given temperature. In liquid medium, the isolates solubilized between 271 and 730 μg ml(-1) of phosphate from tricalcium phosphate. Both the weakest (designated S10) and the strongest solubilizing strain (S06) were further tested for their viability during solubilization. In an assay over the course of 1 week, both strains released their maximum amount of phosphate after 2-4 days. At that later point of time, however, viable cells of isolate S06 were no longer detectable, whereas the weaker strain S10 could be cultured after 1 week in broth. Taking all in vitro observations into account, the usability of the isolates as biofertilizers is critically discussed regarding both the in situ conditions on the tailings pile and the lowered viability due to the excess production of organic acids.     

Molecular mechanism of Endosulfan action in mammals

Endosulfan is a broad-spectrum organochlorine pesticide, speculated to be detrimental to human health in areas of active exposure. However, the molecular insights to its mechanism of action remain poorly understood. In two recent studies, our group investigated the physiological and molecular aspects of endosulfan action using in vitro, ex vivo and in vivo analyses. The results showed that apart from reducing fertility levels in male animals, Endosulfan induced DNA damage that triggers compromised DNA damage response leading to undesirable processing of broken DNA ends. In this review, pesticide use especially of Endosulfan in the Indian scenario is summarized and the importance of our findings, especially the rationalized use of pesticides in the future, is emphasized.